The overall objectives include the following items: 1. Establish the molecular mechanism for the activation of crystalline pyruvate oxidase by phospholipids. 2. Compare phospholipid activated oxidase with trypsin activated oxidase. 3. Determine the mechanism of activation of endonuclease I by colicin E2. 4. Characterize the active site and the amino acid residues at the heme binding site of chloroperoxidase which are responsible for the P-450 character of this enzyme. 5. Examine the relationship between chloroperoxidase compound I and the P-450 hydroxylating intermediate. In the coming year we propose to initiate studies on the amino acid sequence of chloroperoxidase and hopefully identify those amino acid residues responsible for the P-450 character of this protein. We plan to continue our studies on the mechanism of action of colicin E2 primarily by extending our work on the spheroplast reconstitution system. Previous work has shown colicin E2 and endonuclease I are both required to initiate DNA degradation in spheroplast preparation. In contrast, intact cells of Escherichia coli respond to colicin E2 alone.